Daptomycin is a lipopeptide antibiotic that is effective against many gram-positive bacteria strains, including resistant strains such as Methicillin-resistant Staphylococcus aureus and Methicillin-susceptible Staphylococcus aureus. Although daptomycin was approved by the Food and Drug Administration in the early 2000s for the treatment of skin infections and left-sided endocarditis, its mechanism of action remains a topic of ongoing debate. Some proposed mechanisms include membrane depolarization, reorganization, and lipid extrusion, among others. While these mechanisms may be linked to the result of cell death, they do not explain the actual cause of death.

One of the primary objectives of the report is to study the interaction between daptomycin and lipids which are present in the...[ Read more ]

Daptomycin is a lipopeptide antibiotic that is effective against many gram-positive bacteria strains, including resistant strains such as Methicillin-resistant Staphylococcus aureus and Methicillin-susceptible Staphylococcus aureus. Although daptomycin was approved by the Food and Drug Administration in the early 2000s for the treatment of skin infections and left-sided endocarditis, its mechanism of action remains a topic of ongoing debate. Some proposed mechanisms include membrane depolarization, reorganization, and lipid extrusion, among others. While these mechanisms may be linked to the result of cell death, they do not explain the actual cause of death.

One of the primary objectives of the report is to study the interaction between daptomycin and lipids which are present in the bacterial membrane. We employed fluorescence titration experiments to study daptomycin’s change in fluorescence as it interacts with the membrane lipids. Daptomycin showed increased interaction with anionic lipids, which was highest for phosphatidylglycerol (PG). Daptomycin’s affinity to PG was hypothesized to be arising from PG’s headgroup that sets it apart from other anionic lipids, yet Correlation spectroscopy between daptomycin and PG’s headgroup point towards a weak interaction. The interaction between daptomycin and PG is proposed to be synergistic in nature where both the headgroup of PG and the lipid components play a significant role.

Titration between daptomycin and PG in the sub-micromolar range confirmed a strong affinity. Further, daptomycin undergoes irreversible conformational changes upon binding to PG while conformational changes with other anionic lipids are reversible. Although, complex formation between daptomycin and PG had been proposed before but never isolated. We successfully isolated the complex in vitro and in vivo from Bacillus subtilis cells, which were characterized using Reverse phase high performance liquid chromatography and Mass spectrometry.

Daptomycin’s killing mechanism has been a topic of debate for long with recent interest in understanding the initial binding events as the drug binds to the bacterial membrane. We propose a two-stage uptake mechanism for the same. The first is a fast and reversible binding of the drug to the phospholipid membrane in milliseconds and the second is a slow and irreversible insertion into the membrane in minutes, only specific to PG. Consequently, most of the drug accumulates in bacterial membranes rather than in mammalian cells, where it exerts its bactericidal effects through an unknown mechanism within a short period of time. Further, crystallization of the complex and structural elucidation can help us design better analogue of daptomycin to counter the growing drug resistance.